Since 1996, there has been a dramatic and alarming increase in the isolation of multi-drug resistant (MDR) Gram-negative organisms, such as Klebsiella pneumoniae and Escherichia coli, from patients with bloodstream infections, pneumonias, and intra-abdominal infections. Multi-drug resistance among Gram-negative organisms typically denotes bacteria resistant to three or more classes of antibiotics. The increase in MDR Gram-negative bacteria has now been recognized throughout the world and in a number of states within the United States. These MDR bacteria are resistant to not only the cephalosporins but also to the carbapenems (imipenem, meropenem, ertapenem, and doripenem), which have traditionally been the last line of antimicrobial defense against the cephalosporin-resistant organisms. Given that the carbapenems are not effective against these carbapenem-resistant Enterobacteriaceae (CRE), there are now very few antimicrobial options available. In the absence of any new antibiotics to combat these pathogens, healthcare workers are being forced to use older antibiotics such as colistin, which was used in the 1970's and has significant and serious side effects. Moreover, colistin-resistant bacteria are now being identified, reducing the available armamentarium of antimicrobials to one or zero antibiotics. Thus, new therapeutic strategies are urgently needed to fight these MDR Gram-negative pathogens, which carry a high morbidity and mortality rate.
Much of the focus in the last 15 years has been on resistance in Gram-positive organisms (e.g., methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococcus (VRE)), although Gram-negative bacteria account for a large proportion of nosocomial infections. For example, in New York City, Gram-negative organisms account for at least 8% of nosocomial infections, and at least half of those are MDR, carbapenem-resistant bacteria.
The list of antibiotics available to treat infections with any of these MDR CRE organisms is limited from very few to none. Novel approaches are critically needed for identifying new therapies for treating such pathogens, especially since simply producing newer generation antibiotics of the same class is unlikely to provide much benefit since cross-resistance can rapidly develop.
There is a long felt need in the art for compositions and methods useful for fighting Gram-negative organisms and infections caused by these organisms. The present invention satisfies these needs.
Chemokines are chemotactic cytokines that are important regulators of leukocyte-mediated inflammation and immunity in response to a variety of diseases and infectious processes in the host. Chemokines are a superfamily of homologous 8-10 kDa heparin-binding proteins, originally identified for their role in mediating leukocyte recruitment.
The four major families of chemokine ligands are classified on the basis of a conserved amino acid sequence at their amino terminus, and are designated CXC, CC, C, and CX3C sub-families (where “X” is a nonconserved amino acid residue).
The interferon-inducible (ELR−) CXC chemokines are one of the largest families of chemokines, and each member of this group contains four cysteine residues. Most chemokines are small proteins (8-10 kDa in size), have a net positive charge at neutral pH, and share considerable amino acid sequence homology. Structurally, the defining feature of the CXC chemokine family is a motif of four conserved cysteine residues, the first two of which are separated by a non-conserved amino acid, thus constituting the Cys-X-Cys or ‘CXC’ motif. This family is further subdivided on the basis of the presence or absence of another three amino acid sequence, glutamic acid-leucine-arginine (the ‘ELR’ motif), immediately proximal to the CXC sequence. The ELR-positive (ELR+) CXC chemokines, which include IL-8/CXCL8, are potent neutrophil chemoattractants and promote angiogenesis. Among the ELR-negative (ELR−) CXC chemokines, CXCL9, CXCL10 and CXCL11, are potently induced by both type 1 and type 2 interferons (IFN-α/β and IFN-γ, respectively). These Interferon-inducible (ELR−) CXC chemokines are generated by a variety of cell types (including monocytes, macrophages, lymphocytes, and epithelial cells), and are extremely potent chemoattractants for recruiting mononuclear leukocytes, including activated Th1 CD4 T cells, natural killer (NK) cells, NKT cells, and dendritic cells to sites of inflammation and inhibiting angiogenesis.
The chemokine receptors are a family of related receptors that are expressed on the surface of all leukocytes. The shared receptor for CXCL9, CXCL10, and CXCL11 is CXCR3. Through their interaction with CXCR3, the ligands CXCL9, CXCL10 and CXCL11 are the major recruiters of specific leukocytes, including CD4 T cells, NK cells, and myeloid dendritic cells. Importantly, this chemokine ligand-receptor system is at the core of a positive feedback loop escalating Th1 immunity, whereby cytokines such as interleukin (IL)-12 and IL-18 (released by myeloid accessory cells) activate local NK cells to produce IFN-γ, which then induces generation of CXCL9, CXCL10, and CXCL11, which then recruits CXCR3-expressing cells that act as a further source of IFN-γ, which then induces further production of CXCL9, CXCL10, and CXCL11. Consistent with the importance of these interferon-inducible (ELR−) CXC chemokines in promoting Th1-mediated immunity, CXCR3 and its ligands have been documented to play a critical role in host defense against many micro-organisms, including viruses, Mycobacterium tuberculosis, other bacteria, and protozoa.
Independent of their role in CXCR3-dependent leukocyte recruitment, CXCL9, CXCL10, and CXCL11 have recently been found to display direct antimicrobial properties that resemble those of defensins. These antimicrobial effects were first demonstrated in 2001 against Escherichia coli and Listeria monocytogenes. Subsequently, an increasing number of chemokines have been shown to have antimicrobial activity against various strains of bacteria and fungi, including E. coli, S. aureus, Candida albicans, and Cryptococcus neoformans. 
There is a long felt need in the art for new compositions and methods useful as antimicrobial agents, as well as targets for antimicrobial agents. The present invention satisfies these needs.